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1.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-312807

ABSTRACT

SARS-CoV-2 remains a global threat to human health particularly as escape mutants emerge. There is an unmet need for effective treatments against COVID-19 for which neutralizing single domain antibodies (nanobodies) have significant potential. Their small size and stability mean that nanobodies are compatible with respiratory administration. We report four nanobodies (C5, H3, C1, F2) engineered as homotrimers with pmolar affinity for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Crystal structures show C5 and H3 overlap the ACE2 epitope, whilst C1 and F2 bind to a different epitope. Cryo Electron Microscopy shows C5 binding results in an all down arrangement of the Spike protein. C1, H3 and C5 all neutralize the Victoria strain, and the highly transmissible Alpha (B.1.1.7 first identified in Kent, UK) strain and C1 also neutralizes the Beta (B.1.35, first identified in South Africa). Administration of C5-trimer via the respiratory route showed potent therapeutic efficacy in the Syrian hamster model of COVID-19 and separately effective prophylaxis. The molecule was similarly potent by intraperitoneal injection.

2.
Nat Commun ; 12(1): 5469, 2021 09 22.
Article in English | MEDLINE | ID: covidwho-1434103

ABSTRACT

SARS-CoV-2 remains a global threat to human health particularly as escape mutants emerge. There is an unmet need for effective treatments against COVID-19 for which neutralizing single domain antibodies (nanobodies) have significant potential. Their small size and stability mean that nanobodies are compatible with respiratory administration. We report four nanobodies (C5, H3, C1, F2) engineered as homotrimers with pmolar affinity for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Crystal structures show C5 and H3 overlap the ACE2 epitope, whilst C1 and F2 bind to a different epitope. Cryo Electron Microscopy shows C5 binding results in an all down arrangement of the Spike protein. C1, H3 and C5 all neutralize the Victoria strain, and the highly transmissible Alpha (B.1.1.7 first identified in Kent, UK) strain and C1 also neutralizes the Beta (B.1.35, first identified in South Africa). Administration of C5-trimer via the respiratory route showed potent therapeutic efficacy in the Syrian hamster model of COVID-19 and separately, effective prophylaxis. The molecule was similarly potent by intraperitoneal injection.


Subject(s)
Antibodies, Neutralizing/pharmacology , COVID-19/drug therapy , Single-Domain Antibodies/pharmacology , Spike Glycoprotein, Coronavirus/metabolism , Administration, Intranasal , Animals , Antibodies, Neutralizing/administration & dosage , Antibodies, Neutralizing/genetics , Antibodies, Neutralizing/immunology , Cryoelectron Microscopy , Crystallography, X-Ray , Disease Models, Animal , Dose-Response Relationship, Immunologic , Epitopes/chemistry , Epitopes/metabolism , Female , Male , Mesocricetus , Neutralization Tests , SARS-CoV-2/drug effects , Single-Domain Antibodies/administration & dosage , Single-Domain Antibodies/immunology , Single-Domain Antibodies/metabolism , Spike Glycoprotein, Coronavirus/chemistry
3.
FEBS Lett ; 595(18): 2323-2340, 2021 09.
Article in English | MEDLINE | ID: covidwho-1332924

ABSTRACT

The COVID-19 pandemic, caused by the SARS-CoV-2 coronavirus, has triggered a worldwide health emergency. Here, we show that ferritin-like Dps from hyperthermophilic Sulfolobus islandicus, covalently coupled with SARS-CoV-2 antigens via the SpyCatcher system, forms stable multivalent dodecameric vaccine nanoparticles that remain intact even after lyophilisation. Immunisation experiments in mice demonstrated that the SARS-CoV-2 receptor binding domain (RBD) coupled to Dps (RBD-S-Dps) elicited a higher antibody titre and an enhanced neutralising antibody response compared to monomeric RBD. A single immunisation with RBD-S-Dps completely protected hACE2-expressing mice from serious illness and led to viral clearance from the lungs upon SARS-CoV-2 infection. Our data highlight that multimerised SARS-CoV-2 subunit vaccines are a highly efficacious modality, particularly when combined with an ultra-stable scaffold.


Subject(s)
Angiotensin-Converting Enzyme 2/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , Receptors, Virus/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Bacterial Proteins/chemistry , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/chemistry , DNA-Binding Proteins/chemistry , Ferritins/chemistry , Humans , Immunization , Mice , Nanoparticles , Protein Domains , Protein Multimerization , Spike Glycoprotein, Coronavirus/chemistry , Sulfolobus
4.
Adv Med Educ Pract ; 12: 237-243, 2021.
Article in English | MEDLINE | ID: covidwho-1127886

ABSTRACT

INTRODUCTION: During the COVID-19 pandemic, most educational institutions have opted for online education rather than traditional modes of education to protect their employees and students. Online education has been gaining momentum in almost all countries around the world. This coincides with the recently introduced competency-based medical education in India which has embraced online education. This poses a new challenge for the institutions involved, the instructors or teachers, and the students since they must adapt quickly to the new mode of learning. Online education requires teachers to improve their competency in three major areas; pedagogy, technology, and content knowledge. Some of the challenges include; lack of technological skill, poor time management and lack of infrastructure. As technology rapidly advances, health care education systems must also advance in tandem. To implement the new competency-based system and online education, the institutions and the individuals must realize the importance of online education, identify the barriers and quickly work on solutions for success. METHODS: This review was conducted based on various research papers on the topic of online medical education, the challenges faced by faculty members, and the opinion of students on this dilemma. Search terms included online medical education, COVID19, competency-based medical education. CONCLUSION: This review identified various challenges posed by online education on the current medical curriculum, faced by both faculty members and students, especially under the light of the Competency-Based Undergraduate Curriculum for Indian Graduates. Different solutions were proposed to overcome these challenges.

5.
EMBO J ; 40(6): e105543, 2021 03 15.
Article in English | MEDLINE | ID: covidwho-1084490

ABSTRACT

Influenza A virus (IAV) and SARS-CoV-2 (COVID-19) cause pandemic infections where cytokine storm syndrome and lung inflammation lead to high mortality. Given the high social and economic cost of respiratory viruses, there is an urgent need to understand how the airways defend against virus infection. Here we use mice lacking the WD and linker domains of ATG16L1 to demonstrate that ATG16L1-dependent targeting of LC3 to single-membrane, non-autophagosome compartments - referred to as non-canonical autophagy - protects mice from lethal IAV infection. Mice with systemic loss of non-canonical autophagy are exquisitely sensitive to low-pathogenicity IAV where extensive viral replication throughout the lungs, coupled with cytokine amplification mediated by plasmacytoid dendritic cells, leads to fulminant pneumonia, lung inflammation and high mortality. IAV was controlled within epithelial barriers where non-canonical autophagy reduced IAV fusion with endosomes and activation of interferon signalling. Conditional mouse models and ex vivo analysis showed that protection against IAV infection of lung was independent of phagocytes and other leucocytes. This establishes non-canonical autophagy in airway epithelial cells as a novel innate defence that restricts IAV infection and lethal inflammation at respiratory surfaces.


Subject(s)
Autophagy-Related Proteins/genetics , Influenza A virus/pathogenicity , Microtubule-Associated Proteins/metabolism , Orthomyxoviridae Infections/genetics , Sequence Deletion , Alveolar Epithelial Cells/metabolism , Alveolar Epithelial Cells/virology , Animals , Autophagy , Autophagy-Related Proteins/chemistry , Autophagy-Related Proteins/metabolism , Chick Embryo , Cytokines/metabolism , Dogs , Madin Darby Canine Kidney Cells , Mice , Orthomyxoviridae Infections/immunology , Orthomyxoviridae Infections/mortality , Protein Domains , Virus Replication
6.
Viruses ; 12(10)2020 10 14.
Article in English | MEDLINE | ID: covidwho-905965

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). Sequencing the viral genome as the outbreak progresses is important, particularly in the identification of emerging isolates with different pathogenic potential and to identify whether nucleotide changes in the genome will impair clinical diagnostic tools such as real-time PCR assays. Although single nucleotide polymorphisms and point mutations occur during the replication of coronaviruses, one of the biggest drivers in genetic change is recombination. This can manifest itself in insertions and/or deletions in the viral genome. Therefore, sequencing strategies that underpin molecular epidemiology and inform virus biology in patients should take these factors into account. A long amplicon/read length-based RT-PCR sequencing approach focused on the Oxford Nanopore MinION/GridION platforms was developed to identify and sequence the SARS-CoV-2 genome in samples from patients with or suspected of COVID-19. The protocol, termed Rapid Sequencing Long Amplicons (RSLAs) used random primers to generate cDNA from RNA purified from a sample from a patient, followed by single or multiplex PCRs to generate longer amplicons of the viral genome. The base protocol was used to identify SARS-CoV-2 in a variety of clinical samples and proved sensitive in identifying viral RNA in samples from patients that had been declared negative using other nucleic acid-based assays (false negative). Sequencing the amplicons revealed that a number of patients had a proportion of viral genomes with deletions.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/virology , Pneumonia, Viral/virology , Betacoronavirus/isolation & purification , COVID-19 , COVID-19 Testing , COVID-19 Vaccines , Clinical Laboratory Techniques , Coronavirus Infections/diagnosis , DNA, Complementary/analysis , DNA, Complementary/genetics , DNA, Viral/analysis , DNA, Viral/genetics , Genome, Viral , High-Throughput Nucleotide Sequencing/methods , Humans , Molecular Epidemiology , Multiplex Polymerase Chain Reaction , Pandemics , Pneumonia, Viral/diagnosis , RNA, Viral/analysis , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction , SARS-CoV-2 , Sequence Analysis
7.
Viruses ; 12(10):1164, 2020.
Article in English | MDPI | ID: covidwho-855539

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). Sequencing the viral genome as the outbreak progresses is important, particularly in the identification of emerging isolates with different pathogenic potential and to identify whether nucleotide changes in the genome will impair clinical diagnostic tools such as real-time PCR assays. Although single nucleotide polymorphisms and point mutations occur during the replication of coronaviruses, one of the biggest drivers in genetic change is recombination. This can manifest itself in insertions and/or deletions in the viral genome. Therefore, sequencing strategies that underpin molecular epidemiology and inform virus biology in patients should take these factors into account. A long amplicon/read length-based RT-PCR sequencing approach focused on the Oxford Nanopore MinION/GridION platforms was developed to identify and sequence the SARS-CoV-2 genome in samples from patients with or suspected of COVID-19. The protocol, termed Rapid Sequencing Long Amplicons (RSLAs) used random primers to generate cDNA from RNA purified from a sample from a patient, followed by single or multiplex PCRs to generate longer amplicons of the viral genome. The base protocol was used to identify SARS-CoV-2 in a variety of clinical samples and proved sensitive in identifying viral RNA in samples from patients that had been declared negative using other nucleic acid-based assays (false negative). Sequencing the amplicons revealed that a number of patients had a proportion of viral genomes with deletions.

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